Liguo Wang
University of Washington, USA
Title: Random Spherically Constrained (RSC) single particle cryo-EM for structures of membrane proteins in a membrane environment
Biography
Biography: Liguo Wang
Abstract
Recently, a steadily growing community of researchers has been attracted by the capability of cryo-Electron Microscopy (cryo-EM) to obtain atomic-resolution structures without the need to crystalize their samples. To study membrane protein structures using cryo-EM, membrane proteins are usually extracted from cell membranes and dissolved in detergents. However, both functional and structural studies clearly highlight the importance of a lipid membrane environment to preserve protein integrity and activity. To restore the lipid membrane environment of membrane proteins, I have been developing a platform, called “random spherically constrained” (RSC) single-particle cryo-EM, for both structural and functional studies of membrane proteins. The RSC platform establishes the lipid environment for membrane proteins, and makes it possible, for the first time, to apply desired transmembrane potential to trap voltage-gated ion channels in desired functional states for structural analysis. To confirm the absolute amplitude of the transmembrane potential, the hyperpolarization- activated cyclic nucleotide-gated (HCN2) channel was investigated as a model protein, as it only opens at very negative transmembrane potentials. The results confirmed that a negative potential of -120 mV was successfully established as predicted. The RSC method has been successfully employed to obtain the structures of both the large conductance voltage- and calcium-activated potassium (BK) and HCN2 channels in lipid membranes. This confirmed that the RSC method could be used to manipulate the functional states of other voltage-gated ion channels, which is critical for understanding the mechanism under which the ion channel responds to the transmembrane potential.
